Different platforms have been investigated for the production of recombinant proteins including bacterial, yeast, insect and mammalian cells. The main advantage of mammalian cell expression is that the mammalian cell properly and efficiently recognizes the signals for synthesis, processing and secretion of eukaryotic proteins. A diverse range of biologics produced in mammalian cells has been licensed in recent years for therapeutic, diagnostic and research purposes. These include thrombolytics, cytokines, growth factors, immunoglobulins, plasminogen activators, erythropoietines and vaccines.
NS0 is a mammalian cell line widely used in recombinant genetic engineering. NS0 cells are non-immunoglobulin secreting mouse myeloma cells. These cells were originally derived from immunoglobulin-producing tumor cells (NS1 cells), and hence they are well equipped for producing and secreting proteins. NS1 cells express intracellular light chain, while NS0 cells do not.
NS0 cells can be genetically manipulated to express a gene whose product is of interest and are widely used in the bioprocess industry to produce recombinant proteins. Currently, the Glutamine Synthetase-NS0 (GS-NS0) system is most commonly used for expressing recombinant proteins in NS0 cells. The GS system is based upon the extremely low levels of endogenous glutamine synthetase activity in NS0 cells. Exogenous supply of GS through expression of the gene is therefore used for selecting transfectants in glutamine free medium. NS0 cells are much more commonly used than NS1 cells in the industry for the purpose of producing recombinant polypeptides such as antibodies, although NS1 cells can be used for this purpose as well.
NS1 cells and derivatives, including NS0 cells, are cholesterol auxotrophs (Chen et al., Exp. Cell Res., 163(1): 117-126, 1986; Sato et al., In Vitro Cell Dev Biol 24(12):1223-8, 1988; Sato et al., Mol Biol Med 2(2):121-34, 1984). These cells therefore require cholesterol in the media for successful growth. Cholesterol supply and delivery to cells has been particularly difficult to deal with in the bioprocess industry, raising issues involving solubility, filterability, dispersion, physical stability and container adhesion.
The observed cholesterol dependence of NS1 mouse myeloma cells has advantageously been used as a selection mechanism for hybridoma production. Myoken et al. (In vitro Cellular & Developmental Biology: J. Tissue Culture Assoc., 25(5):477-80, May 1989) reported that hybridoma cells produced by the fusion of an NS1 cell and an antibody-producing cell capable of synthesizing cholesterol could be selected from a mixture of such cells and unfused myeloma parent cells by culturing the cells in a cholesterol-free medium. See also U.S. Pat. No. 5,110,737 (Myoken et al.).
However, for applications involving recombinant protein production, for example, cholesterol auxotrophy remains a significant drawback to the use of NS0 and NS1 cells. A mechanism for creating cholesterol independence in NS0 and/or NS1 cells would greatly enhance their utility in many diverse biological applications.